水稻条纹病毒胁迫下的水稻蛋白质组学

 采用双向电泳联用MOLDI-TOF-TOF质谱对水稻感病品种武育粳3号和抗病品种KT95-418感染水稻条纹病毒（Rice stripe virus,RSV）前后的叶片进行蛋白质组学分析。结果显示,RSV基因组编码的病害特异蛋白（disease specific protein,SP）在武育粳3号中的积累量明显高于KT95-418中。其他25个蛋白经质谱成功鉴定,包括RSV NS2蛋白,寄主中与光合作用、细胞氧化还原状态和离子平衡状态及蛋白的合成、转运与翻译后修饰等相关的蛋白。对这些差异表达的蛋白与水稻感、抗病的作用进行了分析。     

Comparative proteomic analysis of cucumber roots infected by Fusarium oxysporum f. sp. cucumerium Owen

Cucumber Fusarium wilt (CFW), caused by the soil-borne fungus Fusarium oxysporum f. sp. cucumerium, is a serious disease in cucumber (Cucumis sativus) production worldwide. For the efficient control of the pathogenic fungi, a better understanding of its interaction and associated resistance mechanisms at the molecular level is required. Here, we report a comparative proteomics analysis of total root protein isolated from infected cucumber root of susceptible bulk (SB) and resistant bulk (RB) of cucumber generation F2. Two-dimensional gel electrophoresis (2-DE) coupled with MS/MS approaches identified 15 over-accumulated proteins from the RB plants. Identified proteins are mainly involved in defense and stress responses, oxidation reduction, metabolism and transport and other process. These proteins are likely to be a part of resistance-related protein network, playing different roles in cucumber disease resistance. Three vital clues regarding wilt resistance of C. sativus are gained from this study. First, jasmonic acid and redox signaling components were found in response to F. oxysporum infection in resistant plants. Second, the LRR family protein may play an important role in the defense reaction against CFW. Third, biotic and abiotic stress-related proteins were induced by the CFW fungus F. oxysporum, indicating the activation of common stress pathway.     

光温敏雄性不育玉米9417雄穗幼穗期叶片叶绿体蛋白质差异分析

为了研究玉米"两用系"9417雄穗不育的分子机制,以玉米光温敏雄性不育系9417的不育和可育2种状态的植株为材料,利用SDS-PAGE、质谱和生物信息学等技术,对其幼穗分化敏感期叶片中叶绿体差异表达蛋白质组进行分离和鉴定。电泳图谱显示,光温敏雄性不育、可育株在雄穗分化发育敏感时期叶绿体表达蛋白至少存在4条显著的差异条带,其中No.1、No.2是可育株中存在而不育株缺失的条带;No.3、No.4是不育株存在可育株中缺失的条带。质谱分析共鉴定出4种蛋白质:①AOL_s00004g466、②luc7-like protein 3-like、③putative T3SS effector EspN2-2、④ABC transporter ATP-binding protein。可育株中No.1为假定蛋白,No.2蛋白的U1-snRNP成分在RNA的加工和修饰中起介导自身与帽结合蛋白相结合的作用;不育株中No.3蛋白具T3SS效应,No.4蛋白在膜上ATP结合区域具单向转运底物的作用。叶绿体出现差异蛋白质,可能与雄性败育、不育有关。     

蛋白质组学技术在鱼类安全生产中的应用研究进展

鱼类水产品以其独特的营养价值,越来越受到人们的关注,鱼类的安全生产也成为研究的热点。近年来,蛋白质组学技术正被逐渐运用到鱼类安全生产的研究中,尤其是鱼类物种的鉴定、鱼类生鲜食物的理化评价、贮藏加工过程中的品质变化、批次质量差异检验和鱼类产品标准化等方面。鉴于此,本文在介绍蛋白质组学技术的基础上,系统探讨了蛋白质组学技术在鱼类安全生产中的应用研究进展,为鱼类安全生产提供一定借鉴。     

Cross comparison of seminal plasma proteins from cattle and buffalo (Bubalus bubalis)

The objective of this study was to evaluate seminal plasma proteins from cattle and buffalo (Bubalus bubalis), to identify differences between related species. Sixteen buffaloes and 16 cattle between 30 and 60 months of age were used. Semen collection was performed by electroejaculation, followed by macroscopic and microscopic subjective analyses. After analysis, the samples were centrifuged at 800 g for 10 min, and the supernatant (seminal plasma) was recentrifuged at 10,000 g for 30 min at 4°C. The total protein concentration was determined by the Bradford method, and the proteins were digested in solution for mass spectrometry (nLC-MS/MS). Multivariate statistical analysis was used to evaluate the proteomics results by non-hierarchical clustering the considering exponentially modified protein abundance index (emPAI). Principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA) were used for clustering. Proteomics identified 78 proteins, and multivariate analysis showed 4 that were over-expressed in buffaloes (cystatin C, prosaposin, peptide YY and keratin type II cytoskeletal 5) and 9 in cattle (spermadhesin-1, seminal plasma protein PDC-109, ribonuclease 4, metalloproteinase inhibitor 2, acrosin inhibitor 1, seminal ribonuclease, C-type natriuretic peptide, angiogenin-1 and osteopontin). Among the proteins identified in seminal plasma, the C-type natriuretic peptide and metalloproteinase inhibitors were described for the first time in buffaloes. Some protease inhibitors were found over-expressed in buffaloes, and important proteins in seminal plasma of cattle were not identified or were found at lower expression levels in buffaloes, which can contribute to reproductive performance in this species.     

组学技术分析肉鸡胚胎发育过程中肝脏蛋白表达的变化

旨在探讨肉鸡胚胎在发育中后期肝脏蛋白水平的变化。本试验选取120枚重量为（65±0.2）g的罗氏308肉鸡种蛋,分为14胚龄（E14）和出壳1日龄（H1）两组,每组3个重复,每个重复20枚种蛋。孵化至E14胚龄和H1日龄时,采集肝脏组织样本,应用相对和绝对定量的等量异位标签（isobaric tags for relative and absolute quantitation,iTRAQ）结合生物信息学分析技术筛选核心差异蛋白。结果显示,在E14和H1日龄之间共筛选出10个核心差异蛋白,主要促进脂肪酸降解（上调ACOX1、ACSL1、ACSL5、CPT1A和ECI2）和糖异生（上调ALDH3A2、FBP1、FBP2、GPI和PGM2,下调LDHB和ALDH9A1）。试验结果提示,胚胎发育的主要能量供给途径是脂肪酸降解,而不是糖酵解。此外,糖异生增多,促进糖原储存以应对出壳以及营养环境变化。     

基于平行反应监测验证技术分析绵羊胚胎骨骼肌蛋白质组数据

旨在对绵羊胚胎骨骼肌蛋白质组学数据进行分析,为阐明绵羊胚胎骨骼肌生长发育机制奠定基础。前期试验应用串联质谱定量法（tandem mass tag,TMT）对成年中国美利奴绵羊妊娠第85（D85）、105（D105）和135天（D135）的胚胎背最长肌进行蛋白质定量,并对D85/D105、D105/D135和D85/D135 3个比较组进行分析。本试验在此基础上,利用KEGG和平行反应监测（parallel reaction monitoring,PRM）等数据分析方法对3个比较组绵羊胚胎骨骼肌上、下调差异丰度蛋白质进行分析和验证,并对候选蛋白质进行生物信息学分析。本研究对肌纤维成熟分化标志性蛋白肌球蛋白重链（myosin-2 isoform X2,MYH）进行PRM靶向定量验证,结果表明,其变化趋势与绵羊胚胎肌纤维成熟分化趋势相符。本研究对上调差异丰度蛋白和下调差异丰度蛋白的KEGG分析结果表明,D105/D85比较组中上调差异丰度蛋白质显著富集于胰岛素等信号通路,D135/D105和D135/D85比较组中下调差异丰度蛋白质显著富集于DNA复制和蛋白质消化吸收等信号通路。与肌肉发育相关的cAMP依赖性蛋白激酶A催化亚基α（PRKACA）和葡萄糖转运蛋白成员4（GLUT4）蛋白均显著富集于胰岛素信号通路中,生物信息学分析显示,这两个蛋白质的理论分子量分别为121.73和20.64 ku,分别有147、12个带正电荷的氨基酸残基和135、12个带负电荷的氨基酸残基;理论等电点分别为8.81和6.54;亲水性平均系数分别为-0.408和0.811;PRKACA蛋白无N端糖基化位点,有45个磷酸化位点;GLUT4蛋白有1个N端糖基化位点,25个磷酸化位点;PRKACA蛋白三级结构与CAMP-2依赖蛋白激酶a嵌合融合的晶体结构相似度为85%,GLUT4蛋白三级结构与人葡萄糖转运蛋白GLUT1的相似性为78%。本研究表明,PRM定量验证趋势与绵羊胚胎肌纤维成熟分化趋势相符,PRKACA和GLUT4蛋白具有丰富的磷酸化修饰位点,并参与胰岛素信号通路调控,是重要的候选蛋白。     

梨品种‘玉露香’抗火疫病蛋白质组学分析

由解淀粉欧文氏菌Erwinia amylovora引起的火疫病是一种重要的经济病害,严重影响着全球苹果和梨的生产。E. amylovora 被列为我国重要的检疫性有害生物。然而,植物抗火疫病分子机制尚不够明确。本研究鉴定了梨品种‘玉露香’对 E. amylovora 的抗性,开展了抗性相关蛋白质组学分析。嫩枝穿刺接种分析结果表明,‘玉露香’对 E. amylovora 表现出明显早期抗性。采用高通量蛋白组学分析技术,鉴定获得‘玉露香’和感病品种‘库尔勒香梨’的差异表达蛋白192个,其中包括29个抗病和防卫相关蛋白,13个氧化还原相关蛋白以及1个铜离子转运ATP酶蛋白HMA5。抑菌动态分析结果显示,2 mmol/L Cu2+完全抑制E. amylovora 的生长。本研究为进一步揭示梨火疫病抗性分子机制提供了候选基因。     

Comparative proteomic analysis of Orobanche and Phelipanche species inferred from seed proteins

Orobanche and Phelipanche species are root parasitic plants some of which infect important crops, causing extensive damage. In an attempt to unravel molecular systematic in these genera using protein markers, we established the protein profile of twelve species by two-dimensional gel electrophoresis (2-DE). Proteins were detected by Sypro Ruby staining and the gel images were digitalised and analysed. The 2-DE maps contained from 30 to 140 protein spots for the analysed species. A virtual master gel was built with spots present in all species and revealing the presence of 260 protein spots. Only one protein was monomorphic, while 143 were unique to a single species. Proteins were scored for the presence or absence of homologous spots to create a binary matrix. Phylogenetic analysis was carried out using maximum parsimony and distance method. The trees obtained were rooted using Cistanche phelypaea as an out-group. Both analyses showed Orobanche as a monophyletic group. AMOVA analysis revealed phenotypic diversity between species within genera, as well as a considerable level of variation between genera. This is a first step to help in the phylogenetic classification of parasitic plants by means of proteomics.     

巴西橡胶树胶乳蛋白质组学研究综述

巴西橡胶树胶乳高速离心后大致可分为橡胶粒子、C-乳清和黄色体3种组分,这些组分中含有约200种不同的蛋白质。随着蛋白质组学的发展,巴西橡胶树胶乳蛋白质组学的研究已相继展开。本文综述近年来巴西橡胶树胶乳蛋白质组学的研究,提出了其存在的问题并进行了展望。